Transcript Transfusion of HHV-8 by blood transfusion NEJM Sept 2006

Transmission of HHV-8
by blood transfusion
NEJM Sept 2006; 355:1331-38
Katerina Pavenski, MD FRCPC
Transfusion Medicine Resident, McMaster University
TMR Journal Club
January 31, 2007
HHV-8 - Meet the Virus
• Discovered in 1994
• Belongs to g herpervirus
subfamily
• Enveloped virus, with 165
kb linear ds DNA genome
(80 ORFs)
• Encodes a number of
host-derived genes that
play a role in modulation
of immune response,
apoptosis and cell growth
- “molecular piracy”
HHV-8 - Meet the Virus
• Exhibits tropism for epithelial and B-cells
• Thought to be exclusively cell-associated
• Following a primary infection, establishes
latency in cells of lymphoid origin
• Virus can be reactivated upon
immunosuppression
HHV-8 - Laboratory Diagnosis
• Serology
– Latent
• Latent nuclear antigen 1 (ORF 73)
– Lytic
• Capsid protein (ORF 65)
• Membrane-associated glycoprotein K8.1
– Note: serologic tests are not standardized and
results may vary widely between laboratories
• Molecular
– PCR for HHV-8 DNA (oral fluids, blood, semen,
etc.)
HHV-8 - Clinical significance
• Primary infection
– Fever, maculopapular rash x 2-14 days (7% of
immunocompetent children in endemic areas)
(Jenson 2003, Andreoni 2002)
– Fever, arthralgia, lymphadenopathy,
splenomegaly, cytopenia (immunocompromised,
frequency unknown) (Cannon 2003)
HHV-8 - Clinical significance
• Reactivation
– Kaposi sarcoma (KS)
• AIDS-associated, African endemic, classical
(Mediterranean), and transplant-associated
• Risk of KS in seropositive individuals
– Healthy and immunocompetent: very low; renal
transplant recipients: 0.2-5%; HIV-positive 30-50%
(Cannon 2003)
– Primary effusion lymphoma
– Multicentric Castleman disease
– Acute bone marrow failure in transplant
recipients
HHV-8 - Prevention and Treatment
• No vaccine available
• No licensed treatment
– HAART in HIV-positive patients reduces
incidence of AIDS KS
– Prophylaxis with anti-herpes drugs
(foscarnet, ganciclovir, cidofovir) may
protect against development of KS
– Anti-herpes drugs may inhibit viral lytic
replication and inhibit KS progression
HHV-8 - Transmission
• Close, nonsexual contact
– Most common route in Africa
– Presumed to be via shedding of virus
from the oral cavity
– Increases throughout childhood and
reaches a plateau by adolescence
• Sexual contact
– Thought to be the most common route in
North America
• Vertical - uncommon
HHV-8 - Transmission
• Parenteral - Arguments against
– Groups with high risk of blood exposure
have lower rates of KS and HHV-8
seropositivity than MSM (Operskalski
1996, Cannon 2003)
– Operskalski et al 1997
• 14 donor-recipient pairs
• 14 donors were seropositive for HIV-1 and
HHV-8
• 10/14 recipients became infected with HIV but
none became HHV-8 seroconverted even
after 19 months
HHV-8 - Transmission
• Parenteral - Arguments against
– Lefrere et al 1997
• 19 patients transfused with leukoreduced
blood on multiple occasions
• None had detectable HHV-8 DNA by PCR
• No serological tests were done
– Others: Engels et al 1999, Marcelin et al
1998
HHV-8 - Transmission
• Parenteral - Arguments for
– Seropositivity increases with increasing
injection drug use and is significantly
associated with Hepatitis C infection (Cannon
et al 2001)
– HHV-8 can be transmitted by infected allografts
(Parravicini 1997, Regamey 1998, Thaunat
2006)
– Case reports suggesting association between
KS and blood transfusion (Davis 1983, VelezGarcia 1985, Cockerill 1986, Bendsoe 1990,
Padilla 1990, Aboulafia 1991)
HHV-8 Transmission
• Parenteral - Arguments for
– Infectious HHV-8 was recovered from a U.S.
blood donor (Blackbourn 1997)
– Viral DNA has been detected in blood donors in
Africa (Belec 1998)
– Seroprevalence of HHV-8 has increased with
increasing number of blood transfusions among
patients with sickle cell anemia in Uganda
(Mbulaiteye 2003)
HHV-8 - Transmission
• Parenteral - Arguments for
– Dollard et al 2005 FACTS (Frequency of
Agents Communicable by Transfusion Study)
• 406 CV surgical patients in Baltimore, 1986-1990
• Change in HHV-8 serostatus measured by lytic
antigen immunofluorescence assay
• 2/284 seronegative patients who have received
transfusions seroconverted
– Both received >10U of non-leukoreduced pRBC
– Linked donor specimens not analyzed
• 0 seronegative patients who did not receive a
transfusion seroconverted
HHV-8 - Seroprevalence
• USA (Cannon et al 2004):
– Gay men 30-50%
– Injection drug users/high risk
heterosexual practices 10-20%
– Blood donors 3%
• Sub-Saharan Africa
– General population 50%
Study
• Research question
– Is risk of HHV-8 seroconversion higher
among recipients of seropositive blood
than among those who receive
seronegative blood?
• Design
– Prospective observational cohort study
Population
• Blood donors:
– Volunteers who donated to the national
blood transfusion service in central
Uganda between November 2000 and
September 2001
– Blood transfused as whole or separated
into RBC and plasma without
leukoreduction
Population
• Transfusion recipients (TR):
– Patients at Mulago Hospital, Kampala
treated between December 2000 and
October 2001
– Inclusion criteria:
• Pretransfusion specimen available and could
be linked to a particular blood unit
• Received a blood transfusion
– Exclusion criteria:
• Previous transfusion within past 6 months
Methods
• Donors
– At the time of donation samples for HHV-8
serology collected
• Transfusion recipients
– Enrollment
• Data: sex, number of children in household, HIV status,
hemoglobin concentration, admission diagnosis,
number of transfusions, volume and component of
blood transfused, duration of blood storage
• Sample for HHV-8 serology (at least 10 days after
blood transfusion)
Methods
• Transfusion recipients
– Follow-up
• 1, 2, and 4 weeks post-transfusion and then
monthly for up to 6 months
• At each visit obtained information about
further transfusions and a sample for HHV-8
serology
• Follow-up ended at the time of last visit,
death, seroconversion, or receipt of an
additional transfusion that was either
seropositive or equivocal
Methods
• Transfusion recipients were included in the
analysis if:
– Pretransfusion specimen was seronegative for
HHV-8 AND at least 2 months of follow-up
completed
– For TR with multiple transfusions during the first 7
days of enrollment, transfusion data was recorded
as a midpoint between 1st and last transfusions
• For seronegative transfusion recipients
– Last two follow-up specimens tested for HHV-8
– If either was positive, then all follow-up specimens
were tested
Methods – Laboratory
• Testing for HHV-8
– Performed in CDC, Atlanta
– Laboratory staff was unaware of the
recipient-donor linkages
– 3 serologic assays (lytic antigens)
• 2 peptide immunoassays based on epitopes
in ORF 65 and K8.1
• Immunofluorescence assay based on lytic
HHV-8 Ag (plasma diluted 1:40 for screen and
1:80 for confirmation)
Methods - Laboratory
• Laboratory results interpretation
– Positive
• Reactivity in 2 or more tests
– Equivocal
• >1 test equivocal with reactivity OR
• Results conflicting or incomplete because of
specimen depletion
Definitions
• Exposed
– TR that received any HHV-8 seropositive blood products
regardless of serological status of additional units
• Unexposed
– Received only HHV-8 negative blood
– Note: patients excluded from analysis if blood received
was serologically equivocal
• Seroconversion
– Two or more consecutive HHV-8 seropositive results
obtained at least 25 days after transfusion
– Date of seroconversion was defined as midpoint between
the last seronegative and the first seropositive visit
Methods – Statistical Analysis
• Excess risk of seroconversion calculated as
the difference between Kaplan-Meier (KM)
survival functions for time to seroconversion
in exposed and unexposed recipients, both
for full follow-up and for the 3-10 week
period after transfusion
• Greenwood’s formula to calculate variance
in excess risk the sum of the variance of KM
estimates
– Confidence intervals calculated by using a
normal approximation
Results – Figure 1
4722
Were not enrolled
101 Received transfusions
from unlinked donors
12 Received transfusions
from donors with
equivocal HHV-8 results
62
Received additional
transfusion from donors with
positive or equivocal HHV-8
results <2 mo after transfusion
6533
1528
1053
Patients with
pretransfusion specimens
Were classified as HHV-8–
seronegative before transfusion
Had sufficient
follow-up
1811
1415
991
Transfusion recipients
enrolled
Had known exposure
status
Were analyzed for
seroconversion
266 Were classified as HHV-8–
seropositive before transfusion
17 Had equivocal results
362
Had insufficient
follow-up
Results – Table 1
Results
• 1811 transfusion recipients enrolled
and followed for average of 4.6 months
• Seroprevalence of HHV-8 at
enrollment
– 36.2% among 1761 linked blood
donations
– 14.5% among recipients before
transfusion; seroprevalence increased
with age
• 820/1811 were excluded from analysis
Results
• 991 patients analyzed
– 425 (42.9%) received seropositive units
– 566 (57.1%) received only seronegative units
• 41/991 (4.1%) seroconverted
– 24 in exposed group and 17 in non-exposed
group
• Exposed patients were significantly more
likely to become infected than unexposed
patients
– Excess risk of seroconversion was 2.8% during
24 weeks of follow-up
Results - Table 2
Results - Figure 2
Results
• From 3-10 weeks post-transfusion, seroconversion
was proportionately more common among exposed
than among unexposed recipients
• Risk of seroconversion was higher among
recipients of seropositive units that had been stored
less than 4 days
• Risk of seroconversion was not associated with the
number of HHV-8 seropositive units transfused, the
volume of blood transfused, the type of blood
component, sex, HIV status or number of children in
the transfusion recipient’s household
Critical Appraisal
• Were there clearly defined groups of
patients, similar in all important ways other
than exposure to HHV-8?
– Yes (see Table 1)
• Was assessment of outcomes either
objective or blinded to exposure to HHV-8?
– Assessment was objective (serological test)
– Laboratory staff performing the HHV-8 testing
was unaware of the recipient-donor linkages
Critical Appraisal
• Was the follow-up of study patients
complete and long enough?
– Follow-up was not complete (362/1415
(26%) lost to follow-up)
– Patients were followed for an average of
4.6 months - probably adequate
• The time from exposure to seroconversion is
not precisely known
Critical Appraisal
• Do the results satisfy “diagnostic tests
for causation”?
– Exposure preceded the onset of outcome
(all seropositive patients and all patients
with history of blood transfusion within
past 6 months excluded)
– Dose-response gradient was NOT
observed (increasing number of
transfusions was not associated with
increased risk of seroconversion)
Critical Appraisal
Is association consistent from study to study?
– YES
• Association between exposure and subsequent
seroconversion supported by some studies (Dollard et al,
etc.)
• Magnitude of risk similar to at least one previous report
(Mbulaiteye 2003)
• Association between exposure and subsequent development
of KS suggested by a few case reports (Moore 2007)
– NO
• Association disputed by other studies (Operskalski 1997,
Lefrere 1997, Engels 1999, etc.)
• Could be explained by smaller sample sizes, low
seroprevalence among donors, presence of leukoreduction
Critical Appraisal
• Does association make biological sense?
• HHV-8 is a cell-associated virus and is likely to
be transmitted by cellular blood products
• The study was meticulously designed to
address potential problems - unstandardized
serological tests, community-acquired
infections, and possibility of passive transfer of
antibody
Critical Appraisal
• Are these valid results of this harm
study important?
exposed
seroconverted
YES
NO
Totals
YES
24
401
425
NO
17
549
566
Critical Appraisal
• Are these valid results of this harm
study important?
Relative risk (RR) = [ a / (a+b) ] / [ c / (c+d) ]
(RR) = [ 24 / 425 ] / [ 17 / 566 ]
(RR) = 1.88
Number needed to harm (NNH) = 1 / [( a / (a+b) ) - ( c / (c+d))]
(NNH) = 1 / [(24 /425) - (17 / 566)]
(NNH) = 38
Critical Appraisal
• Should these valid, potentially important
results change the treatment of our patient?
– Results may not apply to our
circumstances
• HHV-8 seroprevalence among North
American donors is substantially lower (33.5% vs 36%)
• Deferral of donors with risk factors for HHV-8
(IV drug use, high risk sexual behaviour, etc.)
• Universal leukoreduction
• Long RBC storage times
Critical Appraisal
• This study demonstrated association
between exposure and seroconversion but
did not address any clinical outcomes (ex.
subsequent development of KS)
• HHV-8 is most harmful to
immunocompromised patients and arguably
carries little risk to the majority of
transfusion recipients
– Example: patients who have been infected
during transplantation have disease rates of 2564% (Moore 2007)
– Role of selective screening
Critical Appraisal
• Testing limitations
– Serology
• Tests are not standardized
• High throughput serologic assays for HHV-8
do not exist
• Clinical significance of seroreactivity to HHV-8
antigens is questioned in literature
– Molecular
• Reliable techniques exist
• However, most seropositive donors have
been shown to have very low or undetectable
viral load
Critical Appraisal
• Finally, the number of potential
transfusion-transmitted pathogens is
ever increasing and testing for all of
them is not practical/possible
• Restrictive transfusion policy
• Pathogen inactivation, etc.
Conclusion
• HHV-8 can be transmitted by blood
transfusion
• However, currently universal donor
testing in North America is not
warranted
• Further studies are necessary…